DNA replication and repair are complex processes in eukaryotic cells. This application deals with the structure and function of the eukaryotic DNA polymerase-beta and the DNA polymerase- alpha: DNA primase complex that are essential parts of these complex processes. The specific aims are as follow: 1. Molecular cloning of DNA polymerase-beta: cDNA clones of mammalian DNA polymerase-beta will be isolated in order to examine the chromosomal localization, to determine the genomic structure, and to study mRNA turnover during the cell cycle and during DNA repair. Attempts will be made to construct expression plasmids in E. coli that allow overproduction of DNA polymerase-beta so that large quantities of the protein can be prepared for detailed chemical and physical characterization. 2. Molecular cloning of DNA primase peptides from yeast to establish their primary structures and in vivo functions. Molecular cloning of calf thymus DNA polymerase-alpha and DNA primase peptides to establish their primary structures, to examine the chromosomal localization and the genomic organization, and to study the intracellular metabolism of their mRNAs. 3. Biochemical analyses of the calf thymus DNA polymerase- alpha: DNA primase complex including photoaffinity labelling of polypeptides in the complex and search for additional proteins/enzymes required for mammalian DNA replication. 4. To compare the structure of DNA polymerases and DNA primase in normal and transformed cells at protein, RNA and genomic levels.